Smad3-ATF3 signaling mediates TGF-β suppression of genes encoding Phase II detoxifying proteins
Smad3-ATF3 signaling mediates TGF-β suppression of genes encoding Phase II detoxifying proteins
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DOI：
10.1016/j.freeradbiomed.2004.10.033
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发表时间：
2005-02-01
影响因子：
7.4
通讯作者：
Freeman, ML
中科院分区：
文献类型：
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作者：
Bakin, AV;Stourman, NV;Freeman, ML
这项研究提供的证据表明，在乳腺上皮细胞中，多能细胞因子TGF-β 1抑制多个基因的表达，参与II期解毒。GCLC是编码谷氨酸半胱氨酸连接酶催化亚基的基因，谷氨酸半胱氨酸连接酶是谷胱甘肽生物合成中的限速酶，GCLC被用作研究TGF-β抑制II期基因表达的机制的分子替代物。发现TGF-β抑制由人GCLC近端启动子介导的荧光素酶报告基因活性，以及由GCLC抗氧化反应元件ARE 4介导的报告基因活性。TGF-β下调内源性GCLC mRNA和GCLC蛋白的表达。II期基因的TGF-β抑制与细胞谷胱甘肽减少和细胞活性氧增加相关。组成型活性Smad 3E的异位表达足以在没有TGF-β的情况下抑制这两种报告，而显性阴性Smad 3A阻断TGF-β抑制。Smad 3E抑制Nrf 2介导的GCLC报告基因的激活。我们证明，TGF-β增加ATF 3蛋白水平，瞬时过表达Smad 3E。ATF 3的异位表达足以抑制GCLC报告基因活性以及内源性GCLC表达。这些结果表明，Smad 3-ATF 3信号介导的ARE依赖的II期基因表达的TGF-β 1的抑制，并可能提供关键的洞察机制TGF-β 1的功能，在致癌，组织修复和纤维化。(C)2004年爱思唯尔公司All rights reserved.
This study provides evidence that in mammary epithelial cells the pluripotent cytokine TGF-beta1 repressed expression of multiple gene, involved in Phase II detoxification. GCLC, the gene that encodes the catalytic subunit of the enzyme glutamate cysteine ligase, the rate-limiting enzyme in the biosynthesis of glutathione, was used as a molecular surrogate for investigating the mechanisms by which TGF-beta suppressed Phase II gene expression. TGF-beta was found to suppress luciferase reporter activity mediated by the human GCLC proximal promoter, as well as reporter activity mediated by the GCLC antioxidant response element, ARE4. TGF-beta downregulated expression of endogenous GCLC mRNA and GCLC protein. TGF-beta suppression of the Phase II genes correlated with a decrease in cellular glutathione and an increase in cellular reactive oxygen species. Ectopic expression of constitutively active Smad3E was sufficient to inhibit both reporters in the absence of TGF-beta, whereas dominant negative Smad3A blocked TGF-beta suppression. Smad3E suppressed Nrf2-mediated activation the GCLC reporter. We demonstrate that TGF-beta increased ATF3 protein levels, as did transient overexpresssion of Smad3E. Ectopic expression of ATF3 was sufficient to suppress the GCLC reporter activity, as well as endogenous GCLC expression. These results demonstrate that Smad3-ATF3 signaling mediates TGF-beta1 repression of ARE-dependent Phase II gene expression and potentially provide critical insight into mechanisms underlying TGF-beta1 function in carcinogenesis, tissue repair, and fibrosis. (C) 2004 Elsevier Inc. All rights reserved.